IFT88 CRISPR Activation Plasmid (h): sc-404089-ACT

Human IFT88 encodes a core component of the intraflagellar transport (IFT) complex B that is required for primary cilium assembly, maintenance, and anterograde trafficking along axonemal microtubules. By enabling ciliary protein import and turnover, IFT88 supports key sensory and developmental signaling pathways coordinated at the cilium, including Hedgehog and other receptor-mediated cascades that influence cell cycle control and differentiation. Disruption of IFT88-dependent ciliogenesis perturbs ciliary signaling dynamics and cellular polarity, processes broadly implicated in ciliopathies and related disorders affecting tissue morphogenesis and homeostasis. As a central node in cilia biology, IFT88 is frequently studied to connect trafficking defects with pathway misregulation and phenotypes such as altered proliferation, migration, or epithelial organization.

IFT88 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous IFT88 expression without altering the underlying DNA sequence.

IFT88 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the IFT88 locus in human cell lines. The system is built around a catalytically inactive Cas9 (dCas9) carrying two inactivating mutations (D10A and N863A) that eliminate nuclease activity while preserving DNA binding. This dCas9 is fused to VP64, a potent transcriptional activator, and is co-expressed with a blasticidin resistance gene for selection. The second plasmid encodes the MS2-p65-HSF1 fusion protein, a secondary activator complex that works in concert with dCas9-VP64, alongside a hygromycin resistance gene. The third plasmid encodes a target-specific 20 nt sgRNA fused to two MS2 RNA aptamers that recruit the MS2-p65-HSF1 complex to the activation site, accompanied by a puromycin resistance gene. The three plasmids are delivered at a 1:1:1 mass ratio for balanced expression of all system components.

Once assembled at the target locus, the SAM complex binds within approximately 200 bp upstream of the IFT88 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous IFT88 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native IFT88 locus and enabling the study of IFT88-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of IFT88 pathway restoration in tumor cells with silenced or reduced IFT88 expression.

For Research Use Only. Not Intended for Diagnostic or Therapeutic Use.